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Analysis of nuclear protein complexes comprising α-actinin-4 by 2D-electrophoresis and mass spectrometry

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Abstract

Actin-binding protein α-actinin-4 is a member of the spectrin superfamily; it is located in both the cytoplasm and nucleus. However, nuclear functions of α-actinin-4 are still not clear. In this study, we analyzed the composition of nuclear protein complexes associated with α-actinin-4 in A431 cells. Using 2D electrophoresis, we found that about 50 different proteins may be associated with nuclear α-actinin-4. Major proteins of these complexes were analyzed by mass spectrometry and the following proteins have been identified: β-actin, α- and β-tubulins, ribonucleoprotein A2/B1 (regulates splicing and is associated with β-actin), peroxiredoxin-1 (involved in oxidative stress), and glycolytic enzyme D-3-phosphoglycerate dehydrogenase. The detection of these proteins in nuclear complexes with α-actinin-4 may suggest that α-actinin-4 is implicated in transcription and splicing. The presence of β-actin in these complexes was confirmed by tandem mass spectrometry (MALDI-TOF-TOF). The immunoprecipitation of nuclear proteins with antibodies against α-tubulin proved the association of α-actinin-4 with α-tubulin in the protein complex. Nuclear α-actinin-4 consists of a 105-kDa, full-size isoform and two truncated isoforms of 65 and 75 kDa; only truncated isoforms have been found in nuclear complexes with α-tubulin. These data suggest that α-actinin-4 is associated with a number of various nuclear protein complexes that may be able to carry out different functions in the cell nucleus.

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Abbreviations

MS-MS:

method of tandem mass spectrometry

PMF:

peptide mass fingerprint

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Authors and Affiliations

  1. Institute of Cytology, Russian Academy of Sciences, St. Petersburg, Russia

    M. G. Khotin, L. V. Turoverova, A. V. Solovyeva, V. Yu. Aksenova, G. P. Pinaev & D. G. Tentler

  2. Institute for Analytical Instrumentation, Russian Academy of Sciences, St.-Petersburg, Russia

    E. P. Podolskaya & I. A. Krasnov

  3. Linkoping University, Linkoping, Sweden

    K. -E. Magnusson

Authors
  1. M. G. Khotin
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  2. L. V. Turoverova
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  3. E. P. Podolskaya
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  4. I. A. Krasnov
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  5. A. V. Solovyeva
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  6. V. Yu. Aksenova
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  7. K. -E. Magnusson
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  8. G. P. Pinaev
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  9. D. G. Tentler
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Corresponding author

Correspondence to M. G. Khotin.

Additional information

Original Russian Text © M.G. Khotin, L.V. Turoverova, E.P. Podolskaya, I.A. Krasnov, A.V. Solovyeva, V.Yu. Aksenova, K.-E. Magnusson, G.P. Pinaev, D.G. Tentler, 2009, published in Tsitologiya, Vol. 51, No. 8, 2009, pp. 684–690.

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Khotin, M.G., Turoverova, L.V., Podolskaya, E.P. et al. Analysis of nuclear protein complexes comprising α-actinin-4 by 2D-electrophoresis and mass spectrometry. Cell Tiss. Biol. 3, 431–437 (2009). https://doi.org/10.1134/S1990519X09050058

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  • DOI: https://doi.org/10.1134/S1990519X09050058

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